As a step toward understanding the mechanisms of p53 mediated apoptosis and carcinogenesis, we have identified a new putative zinc finger transcription factor, Pw1/ASF-1, and two CED3/ICE-related proteases, NEDD2 and CPP32, whose gene expressions are induced specifically during p53 mediated apoptosis. The main goal of this research project is to investigate the role and regulation of these three gene products in p53-mediated apoptosis. In the first specific aim, significance of the induction of Pw1/ASF-1 during p53 mediated apoptosis will be examined. Biological functions of this protein will be analyzed by expressing the Pw/ASF-1 gene product in mammalian cells. How the Pw1/ASF-1 protein functions as a transcription factor will be explored in specific aim 2. Experiments to identify the DNA binding sites by an in vitro immuno-selection method are proposed. Regulatory mechanism of the Pw1/ASF-1 gene expression will also be investigated by analyzing its promoter and enhancer elements. In the third specific aim, regulation of CED3/ICE related proteases, NEDD2 and CPP32, during p53 mediated apoptosis will be examined. The hypothesis that Induction of protease gene expression can serve as a mechanism to activate CED3/ICE related proteases will be tested. Genetic alteration of p53 is frequently associated with human cancers and p53-mediated apoptosis plays a critical role to prevent carcinogenesis and malignancies, Thus, understanding the mechanism of p53-induced apoptosis represents an important research avenue and may provide a basis for better treatment of human tumors.